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Research in our laboratory focuses on the molecular mechanisms of lymphocyte development in the field of basic medicine. Stem cells of lymphocytes lie in bone marrow and differentiate into B- and T-lineage cells. B-lymphocytes are differentiated and matured in the bone marrow and finally produce antigen-specific antibodies. Precursors of T-lymphocytes move into the thymus and differentiate and mature therein; mature T-lymphocytes then move further to the periphery. In lymphocyte development, the genome-level rearrangement of the lymphocyte antigen receptor (antibody for B-lymphocytes and T-cell receptor for T-lymphocytes) plays an important role; that is, lymphocytes are not developed in the absence of antigen-receptor gene rearrangement. Our laboratory primarily studies the molecular mechanisms of T-lymphocyte development (differentiation, apoptosis, etc.) in the thymus and the transcriptional regulation of recombination-activating genes (RAGs) that are indispensable for antigen-receptor gene rearrangement. Recently, we started to develop a cell chip system for characterizing phenotypes and responses of individual cells in a high-throughput manner. In this study, we are collaborating with people in engineering and manufacturing, and developing a system that detects the cellular responses of about 200,000 lymphocytes at single-cell levels and can pick up single antigen-specific lymphocytes. On the basis of this technique, we founded SC World Inc. (www.scworld.co.jp) and plan to develop antibody based-therapeutics.
www.toyama-mpu.ac.jp/md/immuno/index.html
Antibody cDNA from single B-lymphocytes is recovered by amplifying with a specialized 5′-rapid amplification of cDNA ends (5′-RACE) method. Unlike single-cell reverse transcription PCR (RT-PCR) methods, the strategy is not limited by the particular primer sequences utilized. However, the efficiency of the amplification from single B cells is as high as for the RT-PCR approach. The single cell 5′-RACE method expands the possible target animals from which antibody cDNA can be amplified and extends the target cDNA to the T-cell receptor gene. These capabilities significantly develop the possibility for producing antibody-based therapeutics and T-cell receptor gene-based therapeutics in the future.
Amplification and analysis of cDNA generated from a single cell by 5′-RACE: application to isolation of antibody heavy and light chain variable gene sequences from single B cells, p. 469.
